Using b-cell-targeting antigen igg fusion as tolerogenic protein therapy for treating adverse immune responses

Abstract

The present invention generally relates to antigen-specific tolerogenic protein therapy and the use thereof for treating adverse immune responses, including those associated with autoimmune diseases such as multiple sclerosis (MS) and hemophilia. In particular, the invention involves the application of a B cell-targeting IgG fusion protein as the antigen-specific tolerogenic protein therapy, either alone or in combination with inhibitory antibodies. The fusion protein comprises a B-cell specific targeting module, the constant region of the human IgG4 heavy chain or a fragment thereof; and an antigen.

1 .- 28 . (canceled) 
     
     
         29 . An isolated nucleic acid molecule comprising a nucleotide sequence encoding a fusion protein, wherein said fusion protein comprises an antigen, an IgG heavy chain constant region or a fragment thereof, and a B cell surface targeting molecule. 
     
     
         30 . The isolated nucleic acid molecule of  claim 29 , wherein said IgG heavy chain constant region is a modified human IgG4 heavy chain constant region. 
     
     
         31 . The isolated nucleic acid molecule of  claim 30  wherein said fusion protein does not exhibit B cell depleting efficacy. 
     
     
         32 . The isolated nucleic acid molecule of  claim 31 , wherein said IgG4 heavy chain constant region lacks a hinge region or the CH1 region. 
     
     
         33 . The isolated nucleic acid molecule of  claim 32 , wherein said B cell surface targeting molecule is an anti-CD20 single chain variable fragment or an anti-CD19 single chain variable fragment. 
     
     
         34 . The isolated nucleic acid molecule of  claim 33 , wherein said B cell surface targeting molecule is a humanized anti-CD20 single chain variable fragment comprising an anti-CD20 variable heavy region linked to an anti-CD20 variable light region. 
     
     
         35 . The isolated nucleic acid molecule of  claim 34 , wherein said heavy and light regions are linked via a linker comprising the amino acid sequence (Gly-Gly-Gly-Gly-Ser) 3  (SEQ ID NO: 1). 
     
     
         36 . The isolated nucleic acid molecule of  claim 35 , wherein said protein antigen is selected from the group consisting of myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), proteolipid protein (PLP), Factor VIII C2 domain, Factor VIII A2 domain, or fragments of FVIII domains. 
     
     
         37 . An expression vector comprising the nucleic acid molecule of  claim 36 . 
     
     
         38 . A host cell comprising the expression vector of  claim 37 . 
     
     
         39 . A fusion protein comprising a protein antigen, an IgG heavy chain constant region or a fragment thereof, and a B cell surface targeting molecule. 
     
     
         40 . The fusion protein of  claim 39 , wherein said IgG heavy chain constant region is a modified human IgG4 heavy chain constant region lacking a hinge region or the CH1 region. 
     
     
         41 . The fusion protein of  claim 39 , wherein said B cell surface targeting molecule is an anti-CD20 single chain variable fragment or an anti-CD19 single chain variable fragment. 
     
     
         42 . The fusion protein of  claim 39 , wherein said B cell surface targeting molecule is a humanized anti-CD20 single chain variable fragment comprising an anti-CD20 variable heavy region linked to an anti-CD20 variable light region. 
     
     
         43 . The fusion protein of  claim 39 , wherein said protein antigen is selected from the group consisting of myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG), proteolipid protein (PLP), Factor VIII C2 domain, Factor VIII A2 domain, or fragments of FVIII domains. 
     
     
         44 . A method of inducing tolerogenicity to an endogenous protein in an individual by administering the fusion protein of  claim 39  to said individual. 
     
     
         45 . A method of inducing tolerogenicity to an endogenous protein in an individual by administering the isolated nucleic acid molecule of  claim 29  to said individual. 
     
     
         46 . The method of  claim 44 , further comprising administering a B cell depletion agent. 
     
     
         47 . The method of  claim 46 , wherein said B cell depletion agent reduces the amount of all types of B cells. 
     
     
         48 . The method of  claim 46 , wherein said B cell depletion agent is rituximab or equivalent. 
     
     
         49 . The method of  claim 46 , wherein said B cell depletion agent selectively reduces the amount of follicular B cells and does not reduce the amount of marginal zone B cells or reduces the amount of marginal zone B cells to a lesser extent that follicular B cells. 
     
     
         50 . The method of  claim 49 , wherein said B cell depletion agent is a human equivalent mouse IgG1 isotype anti-CD20 monoclonal antibody. 
     
     
         51 . The method of  claim 44 , wherein said endogenous protein is selected from the group consisting of MBP, MOG, PLP, Factor VIII C2 domain and Factor VIII A2 domain. 
     
     
         52 . The method of  claim 51 , wherein said endogenous protein is MOG and said antigen comprises amino acid residues 35-55 of MOG. 
     
     
         53 . The method of  claim 44 , wherein said individual has been diagnosed with multiple sclerosis. 
     
     
         54 . The method of  claim 44 , wherein said individual has been diagnosed with one of the following diseases, uveitis, type 1 diabetes, arthritis, myasthenia gravis, hemophilia A or B and multiple sclerosis, but also could be used for monogenic enzyme deficiency diseases, such as Pompe's. 
     
     
         55 . The isolated nucleic acid molecule of  claim 29 , wherein the encoded fusion protein comprises, from the N-terminus to the C-terminus, a B cell surface targeting molecule, an antigen, and an IgG heavy chain constant region or a fragment thereof. 
     
     
         56 . The fusion protein of  claim 39 , comprising, from the N-terminus to the C-terminus, a B cell surface targeting molecule, an antigen, and an IgG heavy chain constant region or a fragment thereof. 
     
     
         57 . A host cell comprising the isolated nucleic acid molecule of  claim 36 .